International Journal of
Molecular Sciences Article
Metformin Potentiates the Benefits of Dietary Restraint: A Metabolomic Study Marta Riera-Borrull 1,2 , Anabel García-Heredia 1 ID , Salvador Fernández-Arroyo 1,3 ID , Anna Hernández-Aguilera 1 ID , Noemí Cabré 1 , Elisabet Cuyàs 3,4 , Fedra Luciano-Mateo 1 , Jordi Camps 1 , Javier A. Menendez 3,4 and Jorge Joven 1,5, * 1
2 3 4 5
*
Unitat de Recerca Biomèdica, Hospital Universitari Sant Joan, Institut d’Investigació Sanitària Pere Virgili, Universitat Rovira i Virgili, 43201 Reus, Spain;
[email protected] (M.R.-B.);
[email protected] (A.G.-H.);
[email protected] (S.F.-A.);
[email protected] (A.H.-A.);
[email protected] (N.C.);
[email protected] (F.L.-M.);
[email protected] (J.C.) Centro de Investigaciones Biológicas (CIB-CSIC), 28040 Madrid, Spain Molecular Oncology Group, Girona Biomedical Research Insitiute (IDIBGI), 17190 Girona, Spain;
[email protected] (E.C.);
[email protected] (J.A.M.) ProCURE (Program against Cancer Therapeutic Resistance), Metabolism and Cancer Group, Catalan Institute of Oncology, 17190 Girona, Spain The Campus of International Excellence Southern Catalonia, 43003 Tarragona, Spain Correspondence:
[email protected]; Tel.: +34-977-310-300 (ext. 55409); Fax: +34-977-312-569
Received: 12 September 2017; Accepted: 25 October 2017; Published: 28 October 2017
Abstract: Prevention of the metabolic consequences of a chronic energy-dense/high-fat diet (HFD) represents a public health priority. Metformin is a strong candidate to be incorporated in alternative therapeutic approaches. We used a targeted metabolomic approach to assess changes related to the multi-faceted metabolic disturbances provoked by HFD. We evaluated the protective effects of metformin and explored how pro-inflammatory and metabolic changes respond when mice rendered obese, glucose-intolerant and hyperlipidemic were switched to diet reversal with or without metformin. Mice treated with metformin and diet-reversal showed a dramatically improved protection against HFD-induced hepatic steatosis, a beneficial effect that was accompanied by a lowering of liver-infiltrating pro-inflammatory macrophages and lower release of pro-inflammatory cytokines. Metformin combined with diet reversal promoted effective weight loss along with better glucose control, lowered levels of circulating cholesterol and triglycerides, and reduced adipose tissue content. Our findings underscored the ability of metformin to target the contribution of branched chain amino acids to adipose tissue metabolism while suppressing mitochondrial-dependent biosynthesis in hepatic tissue. The relationship between adipose tissue and liver might provide clinical potential for combining metformin and dietary modifications to protect against the metabolic damage occurring upon excessive dietary fat intake. Keywords: adipose tissue; caloric restriction; energy state; inflammation; liver steatosis; metabolomics
1. Introduction Contemporary high-energy and high-fat diets, coupled with the effect of sedentary lifestyles, are closely related to the convergent epidemics of obesity, insulin resistance, type 2 diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD). These metabolic comorbidities seriously compromise health span and quality of life worldwide [1]. The efficacy of lifestyle modification approaches is low and seriously hampered by relatively unknown cellular metabolic strategies and compensatory pathways induced by reducing fat intake [2,3]. Int. J. Mol. Sci. 2017, 18, 2263; doi:10.3390/ijms18112263
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Severe calorie (energy) restriction (CR) is effective, but it seems impractical outside of research settings. Putative mechanisms are attractive, however, because the adoption of a CR-like lifestyle and the typical diets in industrialized countries operate as extreme ends of the same disease–health metabolic spectrum [4,5]. An alternative strategy, scarcely investigated, might combine lifestyle modification with pharmacotherapy, and in this context efficacy is likely for adenosine monophosphate-activated protein kinase (AMPK) activators [6]. Among available drugs, metformin has been suggested to promote protective effects against metabolic diseases with little or no toxicological significance [7–10]. For example, the combination of metformin and a 70% restriction in calories yielded superior results compared to either treatment alone in diabetic rats [11]. More recently, metformin alone increased energy expenditure and improved liver damage and development of T2D-like state in HFD-fed C57BL6/J mice [12,13]. In mice, taking metformin results in a shift to microbial species with beneficial effects in energy homeostasis [14]. In humans, similar results are likely, and the clinical usefulness of metformin in combination with modest lifestyle modification could be expanded to patients without diabetes with the goal of differentially affecting other medical outcomes [15–19]. How metformin works is not fully understood, but several lines of evidence suggest a critical role in energy homeostasis via a complex picture that includes the effect of nutrients and molecular crosstalk between metabolic organs, especially the liver, gut and adipose tissue [20,21]. Here, we performed an integrated analysis of the metabolic phenotypes occurring upon feeding mice experimental diets with or without metformin treatment using a targeted metabolic approach. We tested the hypothesis that concomitant metformin and simple diet reversal treatment could ameliorate HFD-induced disturbances. 2. Results 2.1. Effects on the Regulation of Body Weight and Food Intake Mice in the HFD group gained weight quicker than chow diet (CD) littermates. Although metformin treatment had no effect on body weight regulation in mice challenged with HFD, it led to a significant reduction in body weight gain in mice on CD from the fourth week of experiment until the end of the follow-up (24 weeks) (Figure 1A). Body weight gain of HFD-fed mice decreased immediately after switching to CD. Remarkably, administration of metformin was significantly more effective than CD alone in reducing body weight gain, particularly after 6–7 weeks of diet reversal when mice fed CD alone began to regain lost body weight (Figure 1B). Despite the gain in weight of HFD mice, the food intake relative to CD-fed-mice was lower. The intake in calories, however, was higher. Data shown in supplementary material indicated differences in energy supply (CD, 3.3 Kcal/g; HFD, 5.7 Kcal/g) per gram of consumed food. The source of these calories was also different. Fat was the main provisor in HFD and carbohydrates in CD. Differences in energy expenditure are unlikely in this model. Metformin treatment had no effect on food consumption between groups (Figure 2A,D).
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Figure 1. Dietary-inducedchanges changes andthe the effect of of metformin on design in in Figure 1. Dietary-induced onweight weightcontrol. control.Overall Overall design Figure 1. Dietary-induced changesand and theeffect effect of metformin metformin on weight control. Overall design in experimental animals. (A) Values obtained in mice upon feeding with experimental diets for 14 experimental animals. (A)(A) Values obtained in mice upon feeding withwith experimental dietsdiets for 14 experimental animals. Values obtained in mice upon feeding experimental forweeks. 14 weeks. HFD,diet; high-fat diet; (B)obtained values obtained 6 weekof period HFD feeding CD, chowCD, diet;chow HFD,diet; high-fat (B) values after a 6 after weekaa period HFD of feeding and shift weeks. CD, chow diet; HFD, high-fat diet; (B) values obtained after 6 week period of HFD feeding and shift to CD. Asterisks denote significant (p < 0.05) changes compared to the respective group. to CD. Asterisks significant (p < 0.05) changes to the respective group. group. and shift to CD.denote Asterisks denote significant (p < 0.05)compared changes compared to the respective
Figure 2. Dietary-induced changes and the effect of metformin on food intake and glucose homeostasis. Figure Dietary-inducedchanges changesand andthe the effect effect of metformin homeostasis. Figure 2. 2. Dietary-induced metforminon onfood foodintake intakeand andglucose glucose homeostasis. (A,D) Cumulative food intake, (B,E) glucose tolerance tests, and (C,F) plasma glucose levels segregated (A,D) Cumulative food intake, (B,E) glucose tolerance tests, and (C,F) plasma glucose levels segregated (A,D) Cumulative food intake, (B,E) glucose tolerance tests, and (C,F) plasma glucose levels segregated by the type of dietary experiment (A–C) CD versus HFD, and (D–F) diet reversal and use of metformin by the type dietary experiment(A–C) (A–C)CD CDversus versus HFD, HFD, and (D–F) diet reversal and use ofof metformin by as theindicated type of of dietary experiment and (D–F) diet reversal and use metformin by the legends. Asterisks denote significant (* p < 0.05; ** p < 0.01) changes. as indicated thelegends. legends.Asterisks Asterisks denote ** p**
